Food Science and Biotechnology

, Volume 23, Issue 6, pp 1805–1813 | Cite as

Biochemical and rheological characterization of a protease from fruits of Withania coagulans with a milk-clotting activity

  • Maryam Beigomi
  • Mohammad Amin Mohammadifar
  • Maryam Hashemi
  • Mohsen Ghods rohani
  • Kalaiselvi Senthil
  • Mohharam Valizadeh
Research Article
First Online:
Received:
Revised:
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Abstract

A protease from Withania coagulans fruits was evaluated for milk-clotting properties. The optimum temperature and pH for enzyme activity were 70°C and 4, respectively. The protease also exhibited an excellent thermal stability at 60°C for 30 min. Fractional precipitation using ammonium sulfate showed that the 40–50% fraction (F5) possessed the highest milk-clotting activity. The F5 fraction showed a Mw band of 66 kDa by SDS-PAGE. Gel formation was monitored using low amplitude oscillatory rheology at different temperatures. An Arrhenius plot was used to describe the temperature dependence of the gelation rate parameter. Time sweep testing showed that an increase in temperature was accompanied by a decrease in the gelation onset time, a higher gel formation rate, and higher values for final gel strength. W.coagulans fruits have potential for use as a rennet substitute.

Keywords

Withania coagulans rheology milk-clotting extraction molecular weight 
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References

  1. 1.
    Hashim MM, Mingsheng D, Iqbal MF, Xiaohong C. Ginger rhizome as a potential source of milk coagulating cysteine protease. Phytochemistry 72: 458–464 (2011)CrossRefGoogle Scholar
  2. 2.
    Tavaria FK, Sousa MJ, Malcata FX. Storage and lyophilization effects of extracts of “Cynara cardunculus” on the degradation of ovine and caprine caseins. Food Chem. 72: 79–88 (2001)CrossRefGoogle Scholar
  3. 3.
    Roseiro LB, Barbosa M, Ames JM, Wilbey RA. Cheesemaking with vegetable coagulants-the use of Cynara L. for the production of ovine milk cheeses. Int. J. Dairy Techol. 56: 76–85 (2003)CrossRefGoogle Scholar
  4. 4.
    Piero AR Lo, Puglisi I, Petrone G. Characterization of “Lettucine”, a serine-like protease from Lactuca sativa leaves, as a novel enzyme for milk clotting. J. Agr. Food. Chem. 50: 2439–2443 (2002)CrossRefGoogle Scholar
  5. 5.
    Oner MD, Akar B. Separation of the proteolytic enzyme from fig tree latex and its utilization in gaziantep cheese production. LWTFood. Sci. Technol. 26: 318–321 (1993)Google Scholar
  6. 6.
    Dinakar P, Mathur M, Roy DD. Differences in proteolytic behaviour in cheddar cheese prepared with calf and vegetable rennet. Indian J. Dairy Sci. 42: 792–796 (1989)Google Scholar
  7. 7.
    Mir BA, Koul S, Soodan AS. Reproductive biology of Withania ashwagandha sp. novo (Solanaceae). Ind. Crop. Prod. 45: 442–446 (2013)CrossRefGoogle Scholar
  8. 8.
    Maurya R, Akanksha, Maurya J, Singh AB, Srivastava AK. Coagulanolide, a withanolide from Withania coagulans fruits and antihyperglycemic activity. Bioorg. Med. Chem. Lett. 18: 6534–6537 (2008)CrossRefGoogle Scholar
  9. 9.
    Anson ML. The estimation of pepsin, trypsin, papain, and cathepsin with hemoglobin. J. Gen. Physiol. 22: 79–89 (1938)CrossRefGoogle Scholar
  10. 10.
    Otani H, Matsumori M, Hosono A. Purification and some properties of a milk clotting protease from the young seeds of Albizia julibrissin. Anim. Sci. Technol. 62: 424–434 (1991)Google Scholar
  11. 11.
    Wang W, Liu QJ, Cui H. Rapid desalting and protein recovery with phenol after ammonium sulfate fractionation. Electrophoresis 28: 2358–2360 (2007)CrossRefGoogle Scholar
  12. 12.
    Bradford MM. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72: 248–254 (1976)CrossRefGoogle Scholar
  13. 13.
    Laemmli UK. Denaturing discontinuous gel electrophoresis. Nature 277: 680–685 (1970)CrossRefGoogle Scholar
  14. 14.
    Castillo M, Lucey J, Payne F. The effect of temperature and inoculum concentration on rheological and light scatter properties of milk coagulated by a combination of bacterial fermentation and chymosin. Cottage cheese-type gels. Int. Dairy J. 16: 131–146 (2006)CrossRefGoogle Scholar
  15. 15.
    Srinivasan M, Lucey J. Effects of added plasmin on the formation and rheological properties of rennet-induced skim milk gels. J. Dairy Sci. 85: 1070–1078 (2002)CrossRefGoogle Scholar
  16. 16.
    Steffe JF. Rheological Methods in Food Process Engineering. 2nd ed. Freeman Press, USA. pp. 158–169 (1996)Google Scholar
  17. 17.
    Gunasekaran S, Ak MM. Cheese rheology and texture. CRC Press. Boca Raton, FL, USA. pp. 22–23 (2003)Google Scholar
  18. 18.
    Lopez MB, Lomholt S, Qvist K. Rheological properties and cutting time of rennet gels. Effect of pH and enzyme concentration. Int. Dairy J. 8: 289–293 (1998)CrossRefGoogle Scholar
  19. 19.
    Esteves CL, Lucey JA, Hyslop DB, Pires E. Effect of gelation temperature on the properties of skim milk gels made from plant coagulants and chymosin. Int. Dairy J. 13: 877–885 (2003)CrossRefGoogle Scholar
  20. 20.
    McMichael AJ. Climate Change and Human Health: Risks and Responses. World Health Organization (WHO), Geneva, Switzerland. pp.18–30 (2003)Google Scholar
  21. 21.
    Reid VJ, Theron LW, Toit Mdu, Divol B. Identification and partial characterization of extracellular aspartic protease genes from Metschnikowia pulcherrima IWBT Y1123 and Candida apicola IWBT Y1384. App. Environ. Microb. 78: 6838–6849 (2012)CrossRefGoogle Scholar
  22. 22.
    Campos R, Guerra R, Aguilar M, Ventura O, Camacho L. Chemical characterization of proteases extracted from wild thistle (Cynara cardunculus). Food Chem. 35: 89–97 (1990)CrossRefGoogle Scholar
  23. 23.
    Ahmed IAM, Morishima I, Babiker EE, Mori N. Characterisation of partially purified milk-clotting enzyme from “Solanum dubium” Fresen seeds. Food Chem. 116: 395–400 (2009)CrossRefGoogle Scholar
  24. 24.
    Purich DL. Enzyme Kinetics: Catalysis & Control: A Reference of Theory and Best-Practice Methods. 1st ed. Elsevier, China. pp. 130–135 (2010)Google Scholar
  25. 25.
    Lamas ME, Barros RM, Balcao VM, Malcata FX. Hydrolysis of whey proteins by proteases extracted from Cynara cardunculus and immobilized onto highly activated supports. Enzyme. Microb. Tech. 28: 642–652 (2001)CrossRefGoogle Scholar
  26. 26.
    Sidrach L, García-Cánovas F, Tudela J, Rodríguez-López JN. Purification of cynarases from artichoke (Cynara scolymus L.): Enzymatic properties of cynarase A. Phytochemistry 66: 41–49 (2005)CrossRefGoogle Scholar
  27. 27.
    Macedo IQ, Faro CJ, Pires EM. Caseinolytic specificity of cardosin, an aspartic protease from the cardoon Cynara cardunculus L.: Action on bovine αs- and β-casein and comparison with chymosin. J. Agr. Food Chem. 41: 1537–1540 (1993)CrossRefGoogle Scholar
  28. 28.
    Guiama V, Libouga D, Ngah E, Mbofung C, Milk-clotting potential of fruit extracts from Solanum esculentum, Solanum macrocarpon L. and Solanum melongena. African. J. Biotech. 9: 3911–3918 (2010)Google Scholar
  29. 29.
    Hooydonk AV, Walstra P. Interpretation of the kinetics of the renneting reaction in milk. Netherlands Milk Dairy J. 41: 19–47 (1987)Google Scholar
  30. 30.
    Silva SV, Malcata FX. Studies pertaining to coagulant and proteolytic activities of plant proteases from Cynara cardunculus. Food Chem. 89: 19–26 (2005)CrossRefGoogle Scholar
  31. 31.
    Van Vliet T. Structure and rheology of gels formed by aggregated protein particles. In: Hydrocolloids. Part 1. Nishinari K (ed). Elsevier Applied Science, Amsterdam, The Netherland. pp. 367–377 (2000)CrossRefGoogle Scholar
  32. 32.
    Hussain I, Grandison AS, Bell AE. Effects of gelation temperature on Mozzarella-type curd made from buffalo and cows’ milk. 1: Rheology and microstructure. Food Chem. 134: 1500–1508 (2012)CrossRefGoogle Scholar
  33. 33.
    Kim BY, Kinsella JE. Rheological changes during slow acid induced gelation of milk by Dgluconoälactone. J. Food Sci. 54: 894–898 (1989)CrossRefGoogle Scholar
  34. 34.
    Lucey J, Singh H. Formation and physical properties of acid milk gels: A review. Food Res. Int. 30: 529–542 (1997)CrossRefGoogle Scholar
  35. 35.
    Zoon P, Vliet T, Walstra P. Rheological properties of rennet-induced skim milk gels. 1. Introduction. Netherlands Milk Dairy J. 42: 249–269 (1988)Google Scholar

Copyright information

© The Korean Society of Food Science and Technology and Springer Science+Business Media Dordrecht 2014

Authors and Affiliations

  • Maryam Beigomi
    • 1
  • Mohammad Amin Mohammadifar
    • 2
  • Maryam Hashemi
    • 3
  • Mohsen Ghods rohani
    • 4
  • Kalaiselvi Senthil
    • 5
  • Mohharam Valizadeh
    • 6
  1. 1.Food and Drug AdministrationZahedan University of Medical ScienceZahedanIran
  2. 2.Department of Food Science and Technology, National Nutrition and Food Technology Research Institute, Faculty of Nutrition Sciences and Food TechnologyShahid Beheshti University of Medical SciencesTehranIran
  3. 3.Microbial Biotechnology & Biosafety DepartmentAgricultural Biotechnology Research Institute of Iran (ABRII)KarajIran
  4. 4.Food Science and Technology GroupInstitute of Scientific Applied Higher Education Jihade-e-AgricultureMashhadIran
  5. 5.Department of BiochemistryBiotechnology and Bioinformatics Avinashilingam University for WomenCoimbatoreIndia
  6. 6.Department of Medicinal and Aromatic PlantHigh Complex Education of SaravanZahedanIran

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